1. Field of the Invention
The invention relates to human coronavirus 229E main proteinase (Mpro) and uses in the development of inhibitors for Severe Acute Respiratory Syndrome (SARS).
2. Description of the Prior Art
Human coronaviruses (HCoV) are major causes of upper respiratory tract illness in humans, in particular, the common cold. To date, only the 229E strain of HCoV has been characterized in detail because it used to be the only isolate that grows efficiently in cell culture. It has recently been shown that a novel coronavirus causes the Severe Acute Respiratory Syndrome (SARS), a disease that is rapidly spreading from its likely origin in Southern China to several countries in other parts of the world. SARS is characterized by high fever, malaise, rigor, headache, non-productive cough or dyspnea and may progress to generalized, interstitial infiltrates in the lung, requiring incubation and mechanical ventilation. The fatality rate among persons with illness meeting the current definition of SARS is around 15% (calculated on outcome, i.e. deaths/deaths+recovered patients). Epidemiological evidence suggests that the transmission of this newly emerging pathogen occurs mainly by face-to-face contact, although airborne transmission cannot be fully excluded. By May 5, 2003, more than 6400 cases of SARS had been diagnosed world-wide, with the numbers still rapidly increasing. At present, no efficacious therapy is available.
Coronaviruses are positive-stranded RNA viruses featuring the largest viral RNA genomes known to date (27-31 kb). The human coronavirus 229E replicase gene, encompassing more than 20,000 nucleotides, encodes two overlapping polyproteins, pp1a (≈450 kD) and pp1ab (≈750 kD) that mediate all the functions required for viral replication and transcription Expression of the COOH-proximal portion of pp1ab requires ribosomal frame shifting. The functional polypeptides are released from the polyproteins by extensive proteolytic processing. This is primarily achieved by the 33.1-kDa HCoV main proteinase (Mpro), also called 3C-like proteinase or 3CLpro, which cleaves the polyprotein at II conserved sites involving mostly Leu-Gln↓(Ser,Ala,Gly) sequences, a process initiated by the enzyme's own autolytic cleavage from pp1a and pp1ab. The functional importance of Mpro in the viral life cycle makes this proteinase an attractive target for the development of drugs directed against SARS and other coronavirus infections.
The design of anticoronaviral drugs directed against the viral main proteinases requires the availability of data on the three-dimensional structures of the target enzymes. In 2002, we determined the crystal structure of the Mpro of transmissible gastroenteritis virus (TGEV), a coronavirus infecting pigs. The structure revealed that coronavirus Mpro consists of three domains, the first two of which together distantly resemble chymotrypsin. However, the catalytic site comprises a Cys-His dyad rather than the Ser-His-Asp triad found in typical chymotrypsin-like serine proteinases.